LPS is an amphiphilic molecule located in the outer leaflet of the outer membrane of Gram-negative bacteria. LPS possesses both endotoxic activity and adjuvant activity. Both properties are based upon its recognition by the host TLR4/MD-2 receptor complex (reviewed in Pålsson-McDermott and O'Neill, 2004; O'Neill, 2006). LPS consists of three distinct structural domains: lipid A, the core, and the O-antigen. Lipid A functions as a hydrophobic membrane anchor and forms the bioactive component of the molecule (Takada and Kotani, 1989). The core region consists of a complex oligosaccharide, which, as compared to the O-antigen, shows only limited structural variability. In some bacteria, e.g., Enterobacteriaceae, the core oligosaccharide (core OS) can be divided into an inner core and an outer core. The outer core primarily consists of pyranosidic hexoses, e.g., D-glucose, D-galactose, and D-glucosamine, whereas the inner core primarily consists of octulosonic acids and heptopyranoses. In the vast majority of Gram-negative bacteria, the core domain is connected to the lipid A domain by a specific carbohydrate, 2-keto-3-deoxyoctulosonic acid (Kdo) (Raetz and Whitfield, 2002). The O-antigen comprises the most variable part of the LPS and confers bacteria serotype specificity. It is composed of repeating sugar subunits of one to eight sugars. Each O-chain can contain up to 50 of these subunits. The O-antigen has been implicated in bacterial immune escape, especially the escape from serum complement-mediated lysis (Raetz and Whitfield, 2002).
In contrast to the LPS of Bordetella bronchiseptica and Bordetella parapertussis, the LPS of Bordetella pertussis never contains an O-antigen domain (Peppler, 1984; Di Fabio et al., 1992). Therefore, B. pertussis LPS is often referred to as lipooligosaccharide. B. pertussis produces two dominant LPS forms, band A and band B LPS (Peppler, 1984). Band B LPS is composed of lipid A and a core oligosaccharide consisting of 9 carbohydrates (Caroff et al., 2000). Addition of a terminal trisaccharide, consisting of N-acetyl glucosamine, 2,3-diacetamido-2,3-dideoxy-mannuronic acid, and 2-acetamido-4-N-methyl-2,4-dideoxy-fucose, to band B LPS forms the LPS referred to as band A.
In Escherichia coli and Salmonella enterica serovar Typhimurium, the core OS biosynthesis gene cluster consists of three operons, designated the gmhD, waaQ, and WaaA operons. The gmhD operon consists of four genes, gmhD and waaFCL, which are involved in the synthesis of the inner core (Schnaitman and Klena, 1993). The gmhD, waaF, and waaC genes encode proteins involved in the biosynthesis and transfer of Heptoses I and II to Kdo2-lipid A (Schnaitman and Klena, 1993), whereas the waaL gene product is a ligase that is involved in the attachment of the O-antigen (MacLachlan et al., 1991). The waaQ operon is the largest of the three operons and encodes proteins that are involved in the biosynthesis of the outer core and in modification/decoration of the core OS. The number and types of genes present within in the waaQ operon differs per strain, which explains the strain-specific differences in core composition (Heinrichs et al., 1998). The waaA operon often encodes only one protein, KdtA. Only in E. coli K-12, an additional non LPS-related open reading frame (ORF) is present (Raetz and Whitfield, 2002). The kdtA gene of Enterobacteriaceae encodes the bifunctional Kdo transferase that adds the two Kdo residues in the Kdo2-lipid A biosynthesis (Clementz and Raetz, 1991).
Although the Bordetella and E. coli core OS show some resemblance, the exact composition and configuration of residues display marked differences. For example, the Bordetella core OS contains only one Kdo residue, instead of the two or three residues that are found in most other Gram-negative bacteria, including E. coli. Recently, this was shown to be due to the functioning of Bordetella KdtA as a monofunctional, rather than as a bifunctional Kdo transferase (Isobe et al., 1999). The enzymes responsible for the synthesis of the remaining portion of the Bordetella core OS are currently unknown and await further identification.
Although its lipid A part is generally seen as the main determinant for the biological activity of LPS through the activation of the TLR4/MD-2 receptor complex, the oligosaccharide region can also play an important role in its interaction with antigen-presenting cells (APCs). Receptors implicated in this type of LPS recognition include the complement receptor CR3 and the scavenger receptor SR-A (van Amersfoort et al., 2003; Plüddemann et al., 2006).
Several Bordetella pertussis vaccines were already used. Introduction of whole-cell pertussis (wP) vaccines in the 1940s and 1950s, and later of acellular pertussis (aP) vaccines in the 1980s and 1990s, led to a gradual decline in pertussis incidence and reduced morbidity and mortality of the disease to low levels. Despite high vaccination coverage, pertussis disease has remained endemic and kept showing a cyclic pattern with peaks in incidence every 2 to 5 years. During the last two decades, several countries, including the Netherlands, have experienced increases in numbers of reported pertussis cases. Interestingly, in some areas, a shift in age distribution has also been observed. Whereas in the pre-vaccination and early vaccine era pertussis cases were predominantly reported in young children, adults and adolescents have accounted for an increasing proportion of the cases in recent years. Several reasons for the re-emergence of reported pertussis have been proposed, including: (1) genetic changes in circulating B. pertussis strains that decrease vaccine efficacy, (2) reduced potency of pertussis vaccines, (3) waning immunity, (4) increased reporting of pertussis cases, and (5) the improved diagnosis of pertussis disease.
Therefore, there is still a need for new vaccines against Bordetella pertussis which does not exhibit all the drawbacks of the existing vaccines.